The present invention relates to the fields of organic chemistry and molecular biology and, more particularly, is directed to a method for synthesizing 4H-chromene derivatives.
Bcl-2 and a family of related proteins regulate apoptosis or programmed cell death and are implicated in a number of human diseases such as cancer. Specifically, Bcl-2 can contribute to neoplastic cell expansion by preventing normal cell turnover caused by physiological cell death mechanisms. High levels of Bcl-2 gene expression are found in a wide variety of human cancers and can lead to tumor cell resistance to conventional chemotherapy and radiotherapy. Synthetic peptides that bind to a functional surface pocket of Bcl-2 have in vitro activity for inducing apoptosis in cell-free systems and in HeLa cells. Furthermore, Bcl-2 binding peptides containing a fatty acid as a cell permeable moiety can induce apoptosis in vitro and have in vivo activity in slowing human myeloid leukemia growth in severe combined immunodeficient mice. These studies suggest that peptides or other small molecules targeted to the Bcl-2 surface pocket could have important clinical applications.
The organic compound HA14-1 (FIG. 1), a 4H-chromene derivative, exhibits binding activity for the surface pocket of Bcl-2 protein (IC50 =9 xcexcM) and induces apoptosis of tumor cells. The discovery of this Bcl-2 binding compound provides a promising lead compound for the development of potential anti-cancer agents and prompted the chemical synthesis of a series of HA14-1 analogs in order to study its structure-activity relationship and increase its potency.
While there are currently two methods for the preparation of 4H-chromene derivatives, both methods have limitations. The first method involves the cyclization of salicylaldehyde derivatives with alkyl cyanoacetates in the presence of ammonium acetate at 5-10xc2x0 C., which produces analogs of HA14-1.
The reaction temperature (5-10xc2x0 C.) is crucial for obtaining the desired products. If the temperature is just slightly raised to 15xc2x0 C., the reaction will fail to give the desired product. In another procedure, aluminum oxide (Al2O3) is used as the catalyst instead of ammonium acetate. However, this procedure is further limited by low yields.
The present invention provides a procedure for the preparation of 4H-chromene derivatives that overcomes the limitations of the current methodology. In one embodiment of the present invention, molecular sieve, more particularly, molecular sieve 3xc3x85, is used as the catalyst. This novel catalyst as disclosed in the present invention, allows the reaction to take place under milder conditions, about 15-300xc2x0 C., and gives higher yields of 4H-chromene derivatives, about 86%.
In the present invention, xe2x80x9c4H-chromene derivativesxe2x80x9d, xe2x80x9cHA14-1 derivativesxe2x80x9d and xe2x80x9cHA14-1 analogsxe2x80x9d are used interchangeably. They include molecules of the formula: 
Within the scope of the present invention, but not being limited thereto, R1 and R2 are CH3, CH2CH3, CH2CHxe2x95x90CH2, CH2Br, CF3, NH2, OH, OCH3, CN, NO2, Cl, Br, F, COOH or COOCH3; and, R3 is selected from the group consisting of hydrogen, CH3, CH2CH3, CH2CH2CH3, CH2CH2CH2CH3, C(CH3)3, CH2Ph or CH2CH2OCH3.
xe2x80x9cstructure-activity relationshipxe2x80x9d as used herein, means the relationship between the structure of a peptide or a molecule and its ability to bind to the functional surface pocket of Bcl-2, thus inhibiting the biological activity of Bcl-2 and inducing apoptosis in cancer cells.